Here we continue studying a new type of regulatory B cells we discovered (Olkhanud et al, Cancer Research, 2011). These cells termed tBregs promote breast cancer metastasis by suppressing antitumor immune responses and inducing the generation of metastasis-promoting Tregs. We also reported that some subsets of human B-CLL are derived from tBregs, thus further reinforcing our original hypothesis that tBregs also exist in humans with cancer (Bodogai et al. Cancer Research, 2013). We have also devised a number of strategies that inactivate tBregs to improve antitumor immune responses (Lee-Chang et al., J. Immunol., 2013; Bodogai et al., Cancer Research, 2013). Because tBregs express low levels of CD20, they cannot be depleted using CD20-targeting antibody rituximab. In fact, the rituximab treatment instead enriches for tBregs and thereby exacerbates metastasis in mice, explaining a recent failure of this strategy in humans with solid tumors. Functionally, tBregs can either directly suppress antitumor T cells. They can also indirectly inhibit T cell activity by inducing the generation of FoxP3+ Tregs and educating MDSCs. The latter feature is most interesting because MDSCs are thought to be the key cancer metastasis-supporting cells. We found that, whereas cancer expands MDSCs with only partially primed activity that is not sufficient to support metastasis, tBregs empower MDSCs with a full regulatory and pro-metastatic functions. We show that tBregs directly activate the regulatory function of both the monocyte and granulocyte subpopulations of MDSC by relying in part on TgfbR1/TgfbR2 signaling. MDSC fully educated in this manner exhibit an increased production of ROS and NO and more efficiently suppress CD4+ and CD8+ T cells, thereby promoting tumor growth and metastasis. Thus, loss of tBregs or Tgfbr deficiency in MDSC is sufficient to disable their suppressive function and to block metastasis. Overall, our data indicate that cancer-induced B cells/B regulatory cells are important regulators of the immune suppressive and pro-metastatic functions of MDSC. This finding has been recently reported (Bodogai et al., Cancer Research, 2015). We also reported that cancer actively converts normal B cells into tBregs by producing and utilizing metabolites of 5-lipoxygenase pathway (5-LOX) and thereby targeting the proliferator-activated receptor alpha (PPARa) signaling in B cells (Wejksza et al., J. Immunology, 2013). Our data also indicate that cancer also uses extracellular vesicles (exosomes) and various soluble factors to support this process. For example, exosomes appear to deliver anti-oxidants to B cells to protect them from the oxidative stress, explaining surprising tolerance of tBregs to reactive oxygen species (ROS). Cancer exosomes were enriched in catalase. Expression of catalase is also highly upregulated in cancer cells that induce tBregs as compared to control cancer cells. At present, we are continuing this study to further elucidate the mechanism of this process. We recently completed the first part of our study on the origin of tBregs. Our data indicate that both in mice and humans, breast cancer uses early B-cell progenitors such as pre-B cells to generate tBregs. Cancer increases Rag, VpreB, IL7R and CD25-expressing B-cell progenitors (CD21- CD23- CD93+ CD25+ Ig- CD19+) in the circulation. To do this, cancer produces thymic stromal lymphopoietin (TSLP), which we show is necessary for expansion and subsequent conversion of CD25Hi pre-B-like cells into tBregs. As such, loss of TSLP expression in cancer cells or TSLPR deficiency in B cells impairs lung metastasis of 4T1 cancer cells. Overall, cancer promotes its metastasis by increasing circulating B-cell progenitors as a source of tBregs. This results we plan to submit for publication in 2018.